This study has been supported by the grant NIHM R01 MH091816 and R21 MH084043 to TDG.

1. Materials 
1. Suspension Box
The tail-suspension test (TST) involves suspending mice above the ground by their tails. At the most basic level, the procedure only requires a suspension bar or shelf ledge, and tape. However, the experimenter should consider the use of a background that provides optimal contrast. Additionally, it is prudent to take steps to prevent mice from observing other animals that are being tested. In our laboratory, we use specially manufactured tail suspension boxes (Four-Hour Day, Baltimore MD), made of plastic with the dimensions (55 height X 60 width X 11.5 cm depth). In order to prevent animals from observing or interacting each other, each mouse is suspended within its own three-walled rectangular compartment (55 height X 15 width X 11.5 cm depth). The mouse is suspended in the middle of this compartment and the width and depth are sufficiently sized so that the mouse cannot make contact with the walls. In this setting, the approximate distance between the mouse's nose and the apparatus floor is 20-25 cm. There are four such identical compartments in the apparatus allowing us to test four mice at a time. An aluminum suspension bar (1 cm. height X 1 cm. width x 60 cm. length), used to suspend the tail of each mouse, is positioned on the top of the box. The dimensions that we use in our laboratory should be regarded as a general reference. For example, the size of individual compartments of the tail-suspension box can be increased if a large outbred mouse strain (e.g. CD-1) is used.
At the bottom of each compartment we place a detachable aluminum tray that collects feces or urine from the animals. We use a dark grey box for albino animals and a cream colored box for mice of other coat colors. This arrangement gives us better contrast and therefore more reliable behavioral scoring of the test.
2. Tape
Tape should adhere securely to both the mouse&#x2019;s tail and the suspension bar, and be strong enough to carry the weight of the mouse being tested (e.g. TimeMed Labeling Systems, Inc, Burr Ridge IL, 1.9 cm width). However, the tape should not be excessively sticky since at the end of the session it will be removed from the tail. We cut 17 cm fragments of tape and place a mark 2 cm from one end. This 2 cm portion was used for attaching the tape to the tail, while the remaining 15 cm was used for suspension of the mouse.
3. Timer



4. Video recording device
A video camera and a tripod (or other support structure) are needed. Since this test usually involves multiple animals being tested at the same time, live scoring will be difficult and is not advised. The video camera should record in high enough resolution to render a quality picture that will be used later for behavioral scoring. Always make certain that there is sufficient recording memory in the camera before starting the test. We use a video camera that records digitally without the use of physical media (i.e. video cassette), allowing for straightforward transfer of videos. 
5. White noise generator
The noise generator should mask intermittent environmental sounds. The use of a noise generator is particularly recommended in laboratory environments where sudden loud noises can be heard that would potentially startle mice. In our experimental room the ambient noise level (without the white noise generator activated) is around 60 dB. The total sound level with the white noise generator activated at the location where the TST is conducted is 70-72 dB. However, it should be noted that these figures are provided as example only, and each laboratory should select the right noise levels according to their unique environment and circumstances.
6. Cleaning supplies
The suspension box should be wiped thoroughly after each session with a sterilization solution (e.g. MB-10, Quip Laboratories Inc., Wilmington, DE, or similar).
7. Climbstoppers (optional: dependent upon strain used)
Mice of some backgrounds, such as C57BL/6, may climb their tails during the test1. Clear hollow cylinders (4 cm length, 1.6 cm outside diameter, 1.3 cm inside diameter, 1.5 grams) that are cut to four cm length (by FourHourDay Inc, Baltimore MD) from polycarbonate tubing (#8585K41, McMaster-Carr, Santa Fe Springs, CA) are placed around the tails of mice to prevent such tail climbing behavior2,3. These devices can be made in any laboratory with common tools and materials. 
2. Behavioral Procedures
The overall experimental design should reflect proper counterbalancing between variables specific to your experiment. For example, in our experiments we try to represent each group equally in every TST session (i.e., if there are four treatment groups, each will be represented in each session). Also, the positions of mice are rotated in a way such that mice from each treatment group are placed in a different position in each session.
<ol>
 <li>Place the camera in position. The camera should be as close as possible in order to obtain the highest possible resolution of the animals.</li>
 <li>Tape fragments that will be used during the session should be cut, marked, and prepared for the session.</li>
 <li>Start the white noise generator, if being used, before the mice are introduced to the testing room. The level of white noise should only be enough to mask external noises. Avoid a high volume and make sure the same level of white noise is used for all animals.</li>
 <li>Bring the animals into the testing room. If the room where the animals reside and the testing room are adjacent to each other with similar ambient conditions, no acclimation period may be necessary. Otherwise, place the animals in the testing room for a period of acclimation (generally at least one hour). Be aware that the other animals placed in the same room can sense olfactory and ultrasonic cues.</li>
 <li>If a strain known to climb their tails, such as C57BL6/J, is being used place Climbstoppers around the tails prior to applying the tape.</li>
 <li>Adhere the tape to the tails of mice. Use the marked end of the tape and make sure the tape sticks to the tail and back on itself. The tape should be applied to the very end of the tail with 2-3 millimeters of tail remaining outside of the tape. </li>
 <li>Once each piece of tape is attached to a mouse tail stick the middle portion of the tape to the inner walls of the cage. This will help prevent tangling of tape pieces to each other. Note that confinement in this fashion may cause stress and that it is important to complete the process of applying the tape to each animal as soon as possible.</li>
 <li>Once all tape is applied, start the recording and identify the session before the mice are suspended.</li>
 <li>Suspend the animals by placing the free end of the tape on the suspension bar or shelf in an order that is counterbalanced between treatment groups. Suspend mice in a way that is does not obstruct the camera view because the entire TST session will be scored and obstructing the camera will lead to the inability to assess behaviors during that time. Check that all tape ends are hanging parallel to the suspension bar or shelf with a similar length of slack for each mouse. </li>
 <li>At the end of the session (which is typically six minutes), return the animals to their homecage and carefully remove the tape from each tail by gently pulling it off. Do not rip the tape from the tail since doing so may cause pain to the mice. </li>
<li>After returning the mice to their colony room discard the fecal boli and urine from the collection trays and wipe the apparatus with a sterilizing solution. </li>
</ol>
3. Behavior Analysis
<ol>
 <li>Generally, the TST is from start to finish six minutes in length. Unlike another widely used antidepressant efficacy procedure, the forced swim test, the whole session is scored. This is due to the general finding that mice tend to manifest immobility earlier in the TST.</li>
 <li>In our laboratory we upload the video files directly from the camera to a PC where the analysis is conducted.</li>
 <li>During the behavioral analysis the time that each mouse spends as mobile is measured. While it is possible to measure the immobility time directly, we have found it easier to detect and measure active movements rather than the lack of such movements. </li>
 <li>The most important aspect of TST behavioral analysis is the consistent identification of movements that are recorded as bona fide mobility. Mice, especially at the beginning of the session, manifest behaviors that are overtly escape related. These include trying to reach the walls of the apparatus and the suspension bar, strong shaking of the body, and movement of the limbs akin to running. These movements clearly constitute mobility. These behaviors then subside and become subtler. In our laboratory small movements that are confined to the front legs but without the involvement of the hind legs are not counted as mobility. Additionally, oscillations and pendulum like swings that are due to the momentum gained during the earlier mobility bouts also are not counted as mobility.</li>
 <li>In our laboratory we use an on-screen stopwatch software for time measurements (Xnote Stopwatch, dnSoft Research Group). Two separate stopwatches are used on the screen. The first stopwatch counts down from 360 seconds and alerts the observer when the behavioral analysis period ends. The second stopwatch &mdash;controlled by the observer&#x2014;measures the time spent mobile. Certain stopwatch software has the ability to assign keys to start and stop functions, so that on-screen the keyboard can control the stopwatch. Instead of a regular keyboard, we use an input device commonly known as a 'gamepad' to control the stopwatches. </li>
 <li>If there is more than one mouse tested and present on the screen it is a good idea to cover the other animals so that their movements will not distract the observer. This can be accomplished by using another program window or by physically covering the other mice on the screen with paper.</li>
 <li>There may be some bias occurring if the total amount of mobility time elapsed for a particular mouse is visible prior to completion of the analysis session. If an on-screen stopwatch is used, we suggest to cover all but the millisecond decimals of the stopwatch. By covering the stopwatch the observer only knows whether the stopwatch is on or off at any point but does not know the total time elapsed and therefore cannot be affected by any bias. Using this approach the observer, while blind to the group assignments of the animals, will not have a general idea of level of mobility in each animal.</li>
 <li>An interobserver reliability test should be conducted for every new observer before beginning to collect data from test animals. In our laboratory each new observer first watches an experienced observer while he or she is scoring. After the new observers gain enough confidence to differentiate mobility from immobility, they then score in real time with the experienced observer watching and pointing out any mistakes. Once this phase is successfully completed, the new observers will analyze a specific set of TST videos that we maintain in our laboratory for training purposes. Only after a high level of interobserver correlation is obtained with the experienced observer does an investigator analyze TST videos from actual experiments. We archive the data from these training analyses to constitute an internal standard for the laboratory. We have observed differences between strains in the manner in which they express mobility (and immobility) behaviors, and mean immobility time between sexes. When a new strain, sex, or genetically modified mouse model is tested in the laboratory it is necessary to again undertake this type of reliability analysis. </li>
</ol>
4. Representative Results
We have used the TST to assess antidepressant-like responses to lithium treatment in various mouse strains (Figure 1)3. Experimental details of this experiment are published in Can et al., 20113. 
<img src="/files/ftp_upload/3769/3769fig1.jpg" alt="Figure 1" /> 
Figure 1. Immobility time in the tail suspension test after chronic lithium administration in five inbred mouse strains. Mice were group housed, four mice per cage. Mice in the lithium treatment group received lithium chow containing 4 g/kg LiCl for three weeks. Control animals received identical food without the lithium chloride. **:p&lt;0.01, ***:p&lt;0.001 denote a significant, unpaired t-test. Data are expressed as mean &plusmn; SEM. n:10-12 animals per group for each strain. (figure reproduced from 3).
In this example, a significant treatment effect of chronic lithium delivered in food is observed only in C57BL/6J and DBA/2J strains. In the other three strains, no statistically significant decrease in immobility time was observed. These data indicate that an antidepressant-like response to chronic lithium treatment is strain dependent as has also been observed with other antidepressant medications4-8.

<ol>
	<li>Mayorga, A. J., Lucki, I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Limitations+on+the+use+of+the+C57BL/6+mouse+in+the+tail+suspension+test.">Limitations on the use of the C57BL/6 mouse in the tail suspension test.</a> Psychopharmacology (Berl). 155, 110-112 (2001).</li><li>Dao, D. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mood+Disorder+Susceptibility+Gene+CACNA1C+Modifies+Mood-Related+Behaviors+in+Mice+and+Interacts+with+Sex+to+Influence+Behavior+in+Mice+and+Diagnosis+in+Humans.">Mood Disorder Susceptibility Gene CACNA1C Modifies Mood-Related Behaviors in Mice and Interacts with Sex to Influence Behavior in Mice and Diagnosis in Humans.</a> Biological Psychiatry. 68, 801-810 (2010).</li><li>Can, A. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Antidepressant-like+responses+to+lithium+in+genetically+diverse+mouse+strains.">Antidepressant-like responses to lithium in genetically diverse mouse strains.</a> Genes, Brain and Behavior. 10, 434-443 (2011).</li><li>Bai, F., Li, X., Clay, M., Lindstrom, T., Skolnick, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Intra-+and+interstrain+differences+in+models+of+"behavioral+despair".">Intra- and interstrain differences in models of "behavioral despair".</a> Pharmacol. Biochem. Behav. 70, 187-192 (2001).</li><li>Cervo, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Genotype-dependent+activity+of+tryptophan+hydroxylase-2+determines+the+response+to+citalopram+in+a+mouse+model+of+depression.">Genotype-dependent activity of tryptophan hydroxylase-2 determines the response to citalopram in a mouse model of depression.</a> J. Neurosci. 25, 8165-8172 (2005).</li><li>David, D. J., Renard, C. E., Jolliet, P., Hascoet, M., Bourin, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Antidepressant-like+effects+in+various+mice+strains+in+the+forced+swimming+test.">Antidepressant-like effects in various mice strains in the forced swimming test.</a> Psychopharmacology (Berl). 166, 373-382 (2003).</li><li>Dulawa, S. C., Holick, K. A., Gundersen, B., Hen, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+of+chronic+fluoxetine+in+animal+models+of+anxiety+and+depression.">Effects of chronic fluoxetine in animal models of anxiety and depression.</a> Neuropsychopharmacology. 29, 1321-1330 (2004).</li><li>Lucki, I., Dalvi, A., Mayorga, A. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sensitivity+to+the+effects+of+pharmacologically+selective+antidepressants+in+different+strains+of+mice.">Sensitivity to the effects of pharmacologically selective antidepressants in different strains of mice.</a> Psychopharmacology (Berl). 155, 315-322 (2001).</li><li>Steru, L., Chermat, R., Thierry, B., Simon, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+tail+suspension+test:+a+new+method+for+screening+antidepressants+in+mice.">The tail suspension test: a new method for screening antidepressants in mice.</a> Psychopharmacology (Berl). 85, 367-370 (1985).</li><li>Can, A., Dao, D. T., Arad, M., Terrillion, C. E., Piantadosi, S. C., Gould, T. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Mouse+Forced+Swim+Test.">The Mouse Forced Swim Test.</a> J. Vis. Exp. 58, 3791-3638 (2012).</li><li>Porsolt, R. D., Bertin, A., Jalfre, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Behavioral+despair+in+mice:+a+primary+screening+test+for+antidepressants.">Behavioral despair in mice: a primary screening test for antidepressants.</a> Arch. Int. Pharmacodyn. Ther. 229, 327-336 (1977).</li><li>Porsolt, R. D., Anton, G., Blavet, N., Jalfre, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Behavioural+despair+in+rats:+a+new+model+sensitive+to+antidepressant+treatments.">Behavioural despair in rats: a new model sensitive to antidepressant treatments.</a> Eur. J. Pharmacol. 47, 379-391 (1978).</li><li>Crowley, J. J., Blendy, J. A., Lucki, I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Strain-dependent+antidepressant-like+effects+of+citalopram+in+the+mouse+tail+suspension+test.">Strain-dependent antidepressant-like effects of citalopram in the mouse tail suspension test.</a> Psychopharmacology (Berl). 183, 257-264 (2005).</li><li>Karolewicz, B., Paul, I. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Group+housing+of+mice+increases+immobility+and+antidepressant+sensitivity+in+the+forced+swim+and+tail+suspension+tests.">Group housing of mice increases immobility and antidepressant sensitivity in the forced swim and tail suspension tests.</a> European Journal of Pharmacology. 415, 197-201 (2001).</li><li>Lad, H. V., Liu, L., Paya-Cano, J. L., Fernandes, C., Schalkwyk, L. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Quantitative+traits+for+the+tail+suspension+test:+automation,+optimization,+and+BXD+RI+mapping.">Quantitative traits for the tail suspension test: automation, optimization, and BXD RI mapping.</a> Mammalian. Genome. 18, 482-491 (2007).</li><li>Liu, X., Gershenfeld, H. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Genetic+differences+in+the+tail-suspension+test+and+its+relationship+to+imipramine+response+among+11+inbred+strains+of+mice.">Genetic differences in the tail-suspension test and its relationship to imipramine response among 11 inbred strains of mice.</a> Biol. Psychiatry. 49, 575-581 (2001).</li><li>Ripoll, N., David, D. J., Dailly, E., Hascoet, M., Bourin, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Antidepressant-like+effects+in+various+mice+strains+in+the+tail+suspension+test..">Antidepressant-like effects in various mice strains in the tail suspension test..</a> Behav. Brain. Res.. 143, 193-200 (2003).</li><li>Mineur, Y. S., Belzung, C., Crusio, W. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+of+unpredictable+chronic+mild+stress+on+anxiety+and+depression-like+behavior+in+mice.">Effects of unpredictable chronic mild stress on anxiety and depression-like behavior in mice.</a> Behav. Brain. Res. 175, 43-50 (2006).</li><li>Thierry, B., Stéru, L., Simon, P., Porsolt, R. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+tail+suspension+test:+Ethical+considerations.">The tail suspension test: Ethical considerations.</a> Psychopharmacology. 90, 284-285 (1986).</li><li>Cryan, J. F., Mombereau, C., Vassout, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+tail+suspension+test+as+a+model+for+assessing+antidepressant+activity:+review+of+pharmacological+and+genetic+studies+in+mice.">The tail suspension test as a model for assessing antidepressant activity: review of pharmacological and genetic studies in mice.</a> Neurosci. Biobehav. Rev. 29, 571-625 (2005).</li><li>O'Leary, O. F., Cryan, J. F., Gould, T. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Mood and Anxiety Related Phenotypes in Mice. 42, 119-137 (2009).</li><li>Detke, M. J., Johnson, J., Lucki, I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Acute+and+Chronic+Antidepressant+Drug+Treatment+in+the+Rat+Forced+Swimming+Test+Model+of+Depression.">Acute and Chronic Antidepressant Drug Treatment in the Rat Forced Swimming Test Model of Depression.</a> Experimental and Clinical Psychopharmacology. 5, 107-112 (1997).</li><li>Rupniak, N. M. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Animal+models+of+depression:+challenges+from+a+drug+development+perspective.">Animal models of depression: challenges from a drug development perspective.</a> Behavioural Pharmacology. 14, 385-390 (2003).</li><li>Strekalova, T., Steinbusch, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Mood and Anxiety related phenotypes in mice: Characterization using behavioral tests. 42, 153-176 (2009).</li><li>Bartolomucci, A., Fuchs, E., Koolhaas, J. M., Ohl, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Mood and Anxiety related phenotypes in mice: Characterization using behavioral tests. 42, 261-275 (2009).</li><li>Samuels, B. A., Hen, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Mood and Anxiety related phenotypes in mice: Characterization using behavioral tests. 63, 107-121 (2011).</li><li>Juszczak, G. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+usage+of+video+analysis+system+for+detection+of+immobility+in+the+tail+suspension+test+in+mice.">The usage of video analysis system for detection of immobility in the tail suspension test in mice.</a> Pharmacology Biochemistry and Behavior. 85, 332-338 (2006).</li><li>Crowley, J. J., Jones, O. '. L. e. a. r. y., F, O., Lucki, I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Automated+tests+for+measuring+the+effects+of+antidepressants+in+mice.">Automated tests for measuring the effects of antidepressants in mice.</a> Pharmacology Biochemistry and Behavior. 78, 269-274 (2004).</li><li>Gould, T. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Involvement+of+AMPA+receptors+in+the+antidepressant-like+effects+of+lithium+in+the+mouse+tail+suspension+test+and+forced+swim+test.">Involvement of AMPA receptors in the antidepressant-like effects of lithium in the mouse tail suspension test and forced swim test.</a> Neuropharmacology. 54, 577-587 (2008).</li><li>Gould, T. D., Dao, D. T., Kovacsics, C. E., Gould, T. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Mood and Anxiety related phenotypes in mice: Characterization using behavioral tests. 42, (2009).</li></ol>

Authors declare no conflicts of interest.

Development of the TST by Steru et al.9 was influenced by the previously developed forced swim test10-12. Similar to the forced swim test, in the TST mice are placed in an inescapable but moderately stressful situation. Lack of escape related behavior is considered immobility. Like the forced swim test, the TST is a test best validated for the evaluation of antidepressant efficacy of drugs, but also used to evaluate the effects of environmental, neurobiological, and genetic manipulations<su...

<ul>
	<li>Tape</li>
	<li>Timer</li>
	<li>Video Camera</li>
	<li>White Noise Generator (optional)</li>
	<li>Climbstoppers (optional; depending upon strain used)</li>
</ul>

the tail suspension test

The tail-suspension test is a mouse behavioral test useful in the screening of potential antidepressant drugs, and assessing of other manipulations that are expected to affect depression related behaviors. Mice are suspended by their tails with tape, in such a position that it cannot escape or hold on to nearby surfaces. During this test, typically six minutes in duration, the resulting escape oriented behaviors are quantified. The tail-suspension test is a valuable tool in drug discovery for high-throughput screening of prospective antidepressant compounds. Here, we describe the details required for implementation of this test with additional emphasis on potential problems that may occur and how to avoid them. We also offer a solution to the tail climbing behavior, a common problem that renders this test useless in some mouse strains, such as the widely used C57BL/6. Specifically, we prevent tail climbing behaviors by passing mouse tails through a small plastic cylinder prior to suspension. Finally, we detail how to manually score the behaviors that are manifested in this test.

Watch this Scientific Journal Video about The Tail Suspension Test at JoVE.com

The Tail Suspension Test

The tail-suspension test is validated as an experimental procedure to assess antidepressant efficacy of drug treatments in mice. Mice are suspended by their tails for six minutes and escape-related behaviors are assessed. We describe procedures used in conducting the tail suspension test.

The tail-suspension test is a mouse behavioral test useful in the screening of potential antidepressant drugs, and assessing of other manipulations that ...

Research

JoVE Journal

Neuroscience

79.1K vistas.  University of Maryland School of Medicine. La prueba de la cola de suspensión se valida como un procedimiento experimental para evaluar la eficacia de los tratamientos farmacológicos antidepresivos en ratones. Los ratones son suspendidos por la cola durante seis minutos y escapar de conductas relacionadas con la evaluación son. Se describen los procedimientos utilizados en la realización de la prueba de suspensión de la cola.

Video: The Tail Suspension Test

Acute and Chronic Tactile Sensory Testing after Spinal Cord Injury in Rats

Spinal cord injury (SCI) impairs sensory systems causing allodynia1-8. To identify cellular and molecular causes of allodynia, sensitive and valid sensory testing in rat SCI models is needed. However, until recently, no single testing approach had been validated for SCI so that standardized methods have not been implemented across labs. Additionally, available testing methods could not be implemented acutely or when severe motor impairments existed, preventing studies of the development of SCI-induced allodynia3. Here we present two validated sensory testing methods using von Frey Hair (VFH) monofilaments which quantify changes in tactile sensory thresholds after SCI4-5. One test is the well-established Up-Down test which demonstrates high sensitivity and specificity across different SCI severities when tested chronically5. The other test is a newly-developed dorsal VFH test that can be applied acutely after SCI when allodynia develops, prior to motor recovery4-5. Each VFH monofilament applies a calibrated force when touched to the skin of the hind paw until it bends. In the up-down method, alternating VFHs of higher or lower forces are used on the plantar L5 dermatome to delineate flexor withdrawal thresholds. Successively higher forces are applied until withdrawal occurs then lower force VFHs are used until withdrawal ceases. The tactile threshold reflects the force required to elicit withdrawal in 50% of the stimuli. For the new test, each VFH is applied to the dorsal L5 dermatome of the paw while the rat is supported by the examiner. The VFH stimulation occurs in ascending order of force until at least 2 of 3 applications at a given force produces paw withdrawal. Tactile sensory threshold is the lowest force to elicit withdrawal 66% of the time. Acclimation, testing and scoring procedures are described. Aberrant trials that require a retest and typical trials are defined. Animal use was approved by Ohio State University Animal Care and Use Committee.

We describe two tactile sensory testing methods for acute or chronic periods of spinal cord injury in rats. These validated procedures can detect the development and maintenance of allodynia-like sensations.

Spinal cord injury (SCI) impairs sensory systems causing allodynia1-8. To identify cellular and molecular causes of allodynia, sensitive and valid sensory ...

Medicine

An Instrumented Pull Test to Characterize Postural Responses

Impairment of postural reflexes, termed postural instability, is a common and disabling deficit in Parkinson's disease. To assess postural reflexes, clinicians typically employ the pull test to grade corrective responses to a backward perturbation at the shoulders. However, the pull test is prone to issues with reliability and scaling (score/4). Here, we present an instrumented version of the pull test to more precisely quantify postural responses. Akin to the clinical test, pulls are manually administered except pull force is also recorded. Displacements of the trunk and feet are captured by a semi-portable motion tracking system. Raw data represent distance traveled (in millimeter units), making subsequent interpretation and analysis intuitive. The instrumented pull test also detects variabilities influencing pull test administration, such as pull force, thereby identifying and quantifying potential confounds that can be accounted for by statistical techniques. The instrumented pull test could have application in studies seeking to capture early abnormalities in postural responses, track postural instability over time, and detect responses to therapy.

Impairment of postural reflexes, termed postural instability, is difficult to quantify. Clinical assessments such as the pull test suffer issues with reliability and scaling. Here, we present an instrumented version of the pull test to objectively characterize postural responses.

Impairment of postural reflexes, termed postural instability, is a common and disabling deficit in Parkinson's disease. To assess postural reflexes, ...

Behavior

Computerized Dynamic Posturography for Postural Control Assessment in Patients with Intermittent Claudication

Computerized dynamic posturography with the EquiTest is an objective technique for measuring postural strategies under challenging static and dynamic conditions. As part of a diagnostic assessment, the early detection of postural deficits is important so that appropriate and targeted interventions can be prescribed. The Sensory Organization Test (SOT) on the EquiTest determines an individual&#39;s use of the sensory systems (somatosensory, visual, and vestibular) that are responsible for postural control. Somatosensory and visual input are altered by the calibrated sway-referenced support surface and visual surround, which move in the anterior-posterior direction in response to the individual&#39;s postural sway. This creates a conflicting sensory experience. The Motor Control Test (MCT) challenges postural control by creating unexpected postural disturbances in the form of backwards and forwards translations. The translations are graded in magnitude and the time to recover from the perturbation is computed.
Intermittent claudication, the most common symptom of peripheral arterial disease, is characterized by a cramping pain in the lower limbs and caused by muscle ischemia secondary to reduced blood flow to working muscles during physical exertion. Claudicants often display poor balance, making them susceptible to falls and activity avoidance. The Ankle Brachial Pressure Index (ABPI) is a noninvasive method for indicating the presence of peripheral arterial disease and intermittent claudication, a common symptom in the lower extremities. ABPI is measured as the highest systolic pressure from either the dorsalis pedis or posterior tibial artery divided by the highest brachial artery systolic pressure from either arm. This paper will focus on the use of computerized dynamic posturography in the assessment of balance in claudicants.

Individuals with intermittent claudication exhibit poor balance compared to healthy controls. Computerized dynamic posturography is an objective method for measuring an individual's postural responses to balance disturbances. This provides an objective reflection of the person&#x2019;s ability to respond to situations under which sensory stimuli are altered and unexpected perturbations occur.

Computerized dynamic posturography with the EquiTest is an objective technique for measuring postural strategies under challenging static and dynamic ...

Back Mechanical Sensitivity Assessment in the Rat for Mechanistic Investigation of Chronic Back Pain

Low back pain is the leading cause of disability worldwide, with dramatic personal, economic, and social consequences. To develop novel therapeutics, animal models are needed to examine the mechanisms and effectiveness of novel therapies from a translational perspective. Several rodent models of back pain are used in current investigations. Surprisingly, however, no standardized behavioral test was validated to assess mechanical sensitivity in back pain models. This is critical to confirm that animals with presumed back pain present local hypersensitivity to nociceptive stimuli, and to monitor sensitivity during interventions designed to relieve back pain. The objective of this study is to lay down a simple and accessible test to assess mechanical sensitivity in the back of rats. A test cage was fabricated specifically for this method; length x width x height: 50 x 20 x 7 cm, having a stainless-steel mesh on the top. This test cage allows the application of mechanical stimuli to the back. To perform the test, the back of the animal is shaved in the region of interest, and the test area is marked to repeat the test on different days, as needed. The mechanical threshold is determined with Von Frey filaments applied to the paraspinal muscles, utilizing the up-down method described previously. The positive responses include (1) muscle twitching, (2) arching (back extension), (3) rotation of the neck (4) scratching or licking the back, and (5) escaping. This behavioral test (Back Mechanical Sensitivity (BMS) test) is useful for mechanistic research with rodent models of back pain for the development of therapeutic interventions for the prevention and management of back pain.

To develop novel therapeutic interventions for the prevention and management of back pain, animal models are required to examine the mechanisms and effectiveness of these therapies from a translational perspective. The present protocol describes the BMS test, a standardized method to assess back mechanical sensitivity in the rat.

Low back pain is the leading cause of disability worldwide, with dramatic personal, economic, and social consequences. To develop novel therapeutics, ...

Validating a Murine Model of Angelman Syndrome Using Behavioral Tests

Behavioral Characterization of an Angelman Syndrome Mouse Model

This manuscript describes a battery of behavioral tests available to characterize Angelman syndrome (AS)-like phenotypes in an established murine model of AS. We use the rotarod learning paradigm, detailed gait analysis, and nest building test to detect and characterize animal motor impairments. We test animal emotionality in the open field and elevated plus maze tests, as well as the affect in the tail suspension test. When AS mice are tested in the open field test, the results should be interpreted with care, since motor dysfunctions influence mouse behavior in the maze and alter activity scores.
The reproducibility and effectiveness of the presented behavioral tests has already been validated in several independent Uba3a mouse lines with different knockout variants, establishing this set of tests as an excellent validation tool in AS research. Models with the relevant construct and face validity will warrant further investigations to elucidate the pathophysiology of the disease and grant the development of causal treatments.

Author Spotlight: A Battery of Highly Reproducible Behavioral Tests to Validate an Angelman Syndrome Murine Model 

This manuscript presents a set of highly reproducible behavioral tests to validate an Angelman syndrome mouse model.

This manuscript describes a battery of behavioral tests available to characterize Angelman syndrome (AS)-like phenotypes in an established murine model of ...

An Alternant Method to the Traditional NASA Hindlimb Unloading Model in Mice

The Morey-Holton hindlimb unloading (HU) method is a widely accepted National Aeronautics and Space Administration (NASA) ground-based model for studying disuse-atrophy in rodents 4-6. Our study evaluated an alternant method to the gold-standard Morey-Holton HU tail-traction technique in mice. Fifty-four female mice (4-8 mo.) were HU for 14 days (n=34) or 28 days (n=20). Recovery from HU was assessed after 3 days of normal cage ambulation following HU (n=22). Aged matched mice (n=76) served as weight-bearing controls.
Prior to HU a tail ring was formed with a 2-0 sterile surgical steel wire that was passed through the 5th, 6th, or 7th inter-vertebral disc space and shaped into a ring from which the mice were suspended. Vertebral location for the tail-ring was selected to appropriately balance animal body weight without interfering with defecation.
We determined the success of this novel HU technique by assessing body weight before and after HU, degree of soleus atrophy, and adrenal mass following HU. Body weight of the mice prior to HU (24.3 &plusmn; 2.9g) did not significantly decline immediately after 14d of HU (22.7 &plusmn; 1.9g), 28d of HU (21.3 + 2.1g) or after 3 days recovery (24.0 &plusmn; 1.8g). Soleus muscle mass significantly declined (-39.1%, and -46.6%) following HU for 14 days and 28 days respectively (p&lt;0.001). Following 3 days of recovery soleus mass significantly increased to 74% of control values. Adrenal weights of HU mice were not different compared to control mice.
The success of our novel HU method is evidenced by the maintenance of animal body weight, comparable adrenal gland weights, and soleus atrophy following HU, corresponding to expected literature values 2, 7, 8. The primary advantages of this HU method include: 1) ease of tail examination during suspension; 2) decreased likelihood of cyanotic, inflamed, and/or necrotic tails frequently observed with tail-taping and HU; 3) no possibility of mice chewing the traction tape and coming out of the suspension apparatus; and 4) rapid recovery and normal cage activity immediately after HU.

We developed an alternant hindlimb unloading model in mice. The primary advantage of our hindlimb unloading tail-ring method over the conventional Morey-Holton tail-traction technique is a simple straightforward procedure that minimizes stress upon the animal.

The Morey-Holton hindlimb unloading (HU) method is a widely accepted National Aeronautics and Space Administration (NASA) ground-based model for studying ...

Biology

A Battery of Motor Tests in a Neonatal Mouse Model of Cerebral Palsy

As the sheer number of transgenic mice strains grow and rodent models of pediatric disease increase, there is an expanding need for a comprehensive, standardized battery of neonatal mouse motor tests. These tests can validate injury or disease models, determine treatment efficacy and/or assess motor behaviors in new transgenic strains. This paper presents a series of neonatal motor tests to evaluate general motor function, including ambulation, hindlimb foot angle, surface righting, negative geotaxis, front- and hindlimb suspension, grasping reflex, four limb grip strength and cliff aversion. Mice between the ages of post-natal day 2 to 14 can be used. In addition, these tests can be used for a wide range of neurological and neuromuscular pathologies, including cerebral palsy, hypoxic-ischemic encephalopathy, traumatic brain injury, spinal cord injury, neurodegenerative diseases, and neuromuscular disorders. These tests can also be used to determine the effects of pharmacological agents, as well as other types of therapeutic interventions. In this paper, motor deficits were evaluated in a novel neonatal mouse model of cerebral palsy that combines hypoxia, ischemia and inflammation. Forty-eight hours after injury, five tests out of the nine showed significant motor deficits: ambulation, hindlimb angle, hindlimb suspension, four limb grip strength, and grasping reflex. These tests revealed weakness in the hindlimbs, as well as fine motor skills such as grasping, which are similar to the motor deficits seen in human cerebral palsy patients.

Presented is a concise battery of mouse neonatal motor tests. Using these tests, neonatal motor deficits can be demonstrated in a variety of neonatal motor disorders. By having a standardized set of tests, results from different studies can be compared, allowing for better and accurate reporting between groups.

As the sheer number of transgenic mice strains grow and rodent models of pediatric disease increase, there is an expanding need for a comprehensive, ...

Thoracic Spinal Cord Hemisection Surgery and Open-Field Locomotor Assessment in the Rat

Spinal cord injury (SCI) causes disturbances in motor, sensory, and autonomic function below the level of the lesion. Experimental animal models are valuable tools to understand the neural mechanisms involved in locomotor recovery after SCI and to design therapies for clinical populations. There are several experimental SCI models including contusion, compression, and transection injuries that are used in a wide variety of species. A hemisection involves the unilateral transection of the spinal cord and disrupts all ascending and descending tracts on one side only. Spinal hemisection produces a highly selective and reproducible injury in comparison to contusion or compression techniques that is useful for investigating neural plasticity in spared and damaged pathways associated with functional recovery. We present a detailed step-by-step protocol for performing a thoracic hemisection at the T8 vertebral level in the rat that results in an initial paralysis of the hindlimb on the side of the lesion with graded spontaneous recovery of locomotor function over several weeks. We also provide a locomotor scoring protocol to assess functional recovery in the open-field. The locomotor assessment provides a linear recovery profile and can be performed both early and repeatedly after injury in order to accurately screen animals for appropriate time points in which to conduct more specialized behavioral testing. The hemisection technique presented can be readily adapted to other transection models and species, and the locomotor assessment can be used in a variety of SCI and other injury models to score locomotor function.

The rat thoracic spinal hemisection is a valuable and reproducible model of unilateral spinal cord injury to investigate the neural mechanisms of locomotor recovery and treatment efficacy. This article includes a detailed step-by-step guide to perform the hemisection procedure and to assess locomotor performance in an open-field arena.

Spinal cord injury (SCI) causes disturbances in motor, sensory, and autonomic function below the level of the lesion. Experimental animal models are ...

Development of a Uterosacral Ligament Suspension Rat Model

Pelvic organ prolapse (POP) is a common pelvic floor disorder (PFD) with the potential to significantly impact a woman's quality of life. Approximately 10%-20% of women undergo pelvic floor repair surgery to treat prolapse in the United States. PFD cases result in an overall $26.3 billion annual cost in the United States alone. This multifactorial condition has a negative impact on the quality of life and yet the treatment options have only dwindled in the recent past. One common surgical option is uterosacral ligament suspension (USLS), which is typically performed by affixing the vaginal vault to the uterosacral ligament in the pelvis. This repair has a lower incidence of complications compared to those with mesh augmentation, but is notable for a relatively high failure rate of up to 40%. Considering the lack of standard animal models to study pelvic floor dysfunction, there is an urgent clinical need for innovation in this field with a focus on developing cost-effective and accessible animal models. In this manuscript, we describe a rat model of USLS involving a complete hysterectomy followed by fixation of the remaining vaginal vault to the uterosacral ligament. The goal of this model is to mimic the procedure performed on women to be able to use the model to then investigate reparative strategies that improve the mechanical integrity of the ligament attachment. Importantly, we also describe the development of an in situ tensile testing procedure to characterize interface integrity at chosen time points following surgical intervention. Overall, this model will be a useful tool for future studies that investigate treatment options for POP repair via USLS.

Pelvic organ prolapse affects millions of women worldwide and yet some common surgical interventions have failure rates as high as 40%. The lack of standard animal models to investigate this condition impedes progress. We propose the following protocol as a model for uterosacral ligament suspension and in vivo tensile testing.

Pelvic organ prolapse (POP) is a common pelvic floor disorder (PFD) with the potential to significantly impact a woman's quality of life. Approximately ...

Aerodynamic Performance of a Model Aircraft: The DC-6B

Source: Jose Roberto Moreto and Xiaofeng Liu, Department of Aerospace Engineering, San Diego State University, San Diego, CA
The low-speed wind tunnel is a valuable tool to study aircraft aerodynamic characteristics and evaluate aircraft performance and stability. Using a scale model of a DC-6B aircraft that has a removable tail and a 6-component external aerodynamic force balance, we can measure the lift coefficient (CL), drag coefficient (CD), pitching moment coefficient (CM), and yaw moment coefficient (CN) of the model airplane with and without its tail and evaluate the effect of the tail on aerodynamic efficiency, longitudinal stability and directional stability.
In this demonstration, airplane aerodynamic characteristics and flight performance and stability are analyzed using the aerodynamic force balance measurement method. This method is widely used in aerospace industries and research labs for aircraft and rocket development. Here, a model DC-6B airplane is analyzed at different flow conditions and configurations, and its behavior is analyzed when it is subjected to sudden changes.

Forces at a Non-zero Windspeed in the DC-6B Model Aircraft

Source: Jose Roberto Moreto and Xiaofeng Liu, Department of Aerospace Engineering, San Diego State University, San Diego, CA
The low-speed wind tunnel is ...